This invention relates to nuclear magnetic resonance imaging (MRI) and, more particularly, to extended-linear polymeric contrast agents for magnetic resonance imaging of tumors and methods of synthesizing such agents.
Tumor angiogenesis is the recruitment of new blood vessels by a growing tumor from existing neighboring vessels. This recruitment of new microvasculature is a central process in tumor growth and in the potential for aggressive spreading of the tumor through metastasis. All solid tumors require angiogenesis for growth and metastasis. Thus, the level of angiogenesis is thought to be an important parameter for the staging of tumors. Furthermore, new therapies are being developed which attack the process of angiogenesis for the purpose of attempting to control tumor growth and tumor spread by restricting or eliminating the tumor blood supply. It is therefore of clinical importance to be able to monitor angiogenesis in tumors in a noninvasive manner.
To assess angiogenic activity of tumors, two parameters are of primary importance: vascular volume and vascular permeability. Invasive techniques utilizing tissue staining can be used to assess microvascular development, but the sensitivity of existing staining methods is not high enough and the prognostic value of such methods is not yet well established (N. Weidner, et al., New Eng. J. Med. 324:1-8, 1991). At present there is no single imaging method capable of providing quantitative characterization of tumor angiogenesis.
As for non-invasive methods for assessing the two parameters, the parent application Ser. No. 09/451,719 xe2x80x9cnow U.S. Pat. No. 6,235,264xe2x80x9d, teaches a magnetic resonance imaging method with a type of contrast agent that enables measurement of both vascular volume and vascular permeability with much higher sensitivity than heretofore possible. Such measurement should facilitate independent prognostic assessments of cancer and help in monitoring cancer therapy non-invasively.
When a substance such as living tissue is subjected to a uniform magnetic field (polarizing field B0), individual magnetic moments of the nuclear spins in the tissue attempt to align with this polarizing field along the z axis of a Cartesian coordinate system, but process about the z axis direction in random order at their characteristic Larmor frequency. If the substance, or tissue, is subjected to a magnetic field (excitation field B1) which is in the x-y plane and at a frequency near the Larmor frequency, the net aligned longitudinal magnetization may be rotated, or xe2x80x9ctippedxe2x80x9d, into the x-y plane to produce a net transverse magnetization. A signal is emitted by the excited spins after the excitation signal B1 is terminated. This NMR signal may be received and processed to form an image.
When utilizing NMR signals of this type to produce images, magnetic field gradients (Gx Gy and Gz) are employed. Typically, the region to be imaged is scanned with a series of measurement cycles in which these gradients vary according to the particular localization method being used. The resulting set of received NMR signals is digitized and processed to reconstruct the image using one of many well known reconstruction techniques.
One of the mechanisms employed in MRI to provide contrast in reconstructed images is the T1 relaxation time of the spins. After excitation, a period of time is required for the longitudinal magnetization to fully recover. This period, referred to as the T1 relaxation time, varies in length depending on the particular spin species being imaged. Spin magnetizations with shorter T1 relaxation times appear brighter in MR images acquired using fast, T1 weighted NMR measurement cycles. A number of contrast agents which reduce the T1 relaxation time of neighboring water protons are used as in vivo markers in MR images. The level of signal brightness, i.e., signal enhancement, in T1 weighted images is proportional to the concentration of the agents in the tissue being observed.
In pre-clinical research applications, high-field MRI has been used to assess tumor volume and tumor signal changes in animal models after treatment with tamoxifen, a type of antiangiogenic agent (H. E. Maretzek, et al., Cancer Res., 54:5511-5514, 1994). By using an intravascular contrast agent, albumin-Gd-DTPA, tumor vascular volume and permeability were measured as well as spatial distribution of the neovasculature. In another study using a high polarizing field, tumor growth was followed by using a variety of NMR measurement pulse sequences that allowed the investigators to distinguish microvessels from larger vessels through blood oxygen level dependent effects. Permeability was assessed by noting the time dependent changes in NMR signal when Gd-DTPA was administered to the animal (R. Abramovitch, et al., Cancer Res. 55:1956-1962, 1995).
At lower polarizing fields that are available at clinical sites, Gd-DTPA, an MRI contrast agent approved by the FDA (U.S. Food and Drug Administration) has been used to estimate angiogenic activity of tumors (C. Frouge, et al., Invest. Radiol. 29:1043-1049, 1994). However, this contrast agent is not ideal for characterizing tumor vasculature because it rapidly migrates to the extravascular space before being excreted through the kidneys. The tumor NMR signal measurements become delicate, being based on the dynamics of contrast agent uptake and elimination. Staging of tumors by this approach has been difficult (R. Brasch, et al., Radiology 200:639-649, 1996).
To avoid the delicate dynamic aspects of Gd-DTPA uptake measurements, others have used a macromolecular contrast agent, albuminxe2x80x94Gd-DTPA (F. Demser, et al., Mag. Res. Med. 37:236-242, 1997). In this instance, the elimination process does not play a role in the observed MR signals, so that a much simpler and more reliable signal analysis is possible. Thus, MR signals based on T1 changes (proportional to agent concentration) have provided indications of tumor blood vessel leak rate and tumor blood volume. This then represents an effective imaging method for assessing tumor angiogenesis. A severe drawback to this approach, however, is that this macromolecular agent has associated immune reactions when injected and leads to substantial toxicities. Thus, at present, this contrast agent is unsuitable for clinical applications (T. J. Passe, et al., Radiology 230:593-600, 1997).
In a preferred embodiment of the invention, a contrast agent for use in acquiring MRI images for the purpose of assessing tumor angiogenesis comprises a reptating polymer containing gadolinium. Methods for synthesizing this polymer and linear extended polymeric paramagnetic chelates for use as MRI contrast agents are provided wherein DTPA (diethylene triamine pentaacetic acid) chelator moieties are conjugated to higher than 90% of the monomer residues of the polyamino acid backbone chain. The resulting polymer can be labeled with Gd, since each chelator moiety will hold a Gd ion, and the resulting conformation is of an unfolded, extended linear type, capable of entering small pores and moving around obstacles in the extracellular space of living tissues. Efficient production of these extended polymers is critical for the application of such contrast agents to medical imaging.